LncRNA LINC00466 Promotes the Progression of Breast Cancer via miR-4731-5p/EPHA2 Pathway.

BACKGROUND: Breast Cancer (BC) is a female malignancy with a high mortality rate. Novel diagnostic and prognostic biomarkers are valuable for reducing BC mortality. Our study is designed to undrape the precise role of the LINC00466/miR-4731-5p/EPHA2 axis in BC.

Explore on the Mechanism of miRNA-146a/TAB1 in the Regulation of Cellular Apoptosis and Inflammation in Ulcerative Colitis Based on NF-κB Pathway.

OBJECTIVE: Ulcerative colitis (UC) is a chronic non-specific inflammatory disease of the rectum and colon with unknown etiology. A growing number of evidence suggest that the pathogenesis of UC is related to excessive apoptosis and production of inflammatory cytokines. However, the functions and molecular mechanisms associated with UC remain unclear. MATERIALS AND METHODS: The in vivo and in vitro models of UC were established in this study. MiRNA or gene expression was measured by qRT-PCR assay. ELISA, CCK-8, TUNEL, and flow cytometry assays were applied for analyzing cellular functions. The interactions between miR-146a and TAB1 were verified by luciferase reporter and miRNA pull-down assays. RESULTS: MiR-146a was obviously increased in UC patients, DSS-induced colitis mice, and TNF-ɑ-induced YAMC cells, when compared to the corresponding controls. MiR- 146a knockdown inhibited the inflammatory response and apoptosis in DSS-induced colitis mice and TNF-ɑ-induced YAMC cells. Mechanistically, we found that TAB1 was the target of miR-146a and miR-146a knockdown suppressed the activation of NF-κB pathway in UC. More importantly, TAB1 could overturn the inhibitory effect of antagomiR-146a on cell apoptosis and inflammation in UC. CONCLUSION: MiR-146a knockdown inhibited cell apoptosis and inflammation via targeting TAB1 and suppressing NF-κB pathway, suggesting that miR-146a may be a new therapeutic target for UC treatment.

LINC02086 promotes cell viability and inhibits cell apoptosis in breast cancer by sponging miR-6757-5p and up-regulating EPHA2.

BACKGROUND: Long non-coding RNAs (lncRNAs) are critical regulators in the initiation and progression of breast cancer. Our study aims to characterize the functions of LINC02086 which few published in breast cancer and decipher the downstream molecular mechanisms. METHODS: LINC02086 expression is tested in RNA-seq data from GEPIA database, tumor tissue samples from hospital patients and breast cancer cell lines. LINC02086 was silenced or overexpressed by lenti-virus-mediated shRNAs, or pLVX-Puro plasmids. Luciferase reporter assay and RNA pull-down assay were applied to study interactions between LINC02086, miR-6757-5p and ephrin type-A receptor 2 (EPHA2). LINC02086-silencing MCF-7 cells were injected into mice to establish xenograft animal models. RESULTS: Using RNA-seq data, tumor tissue samples and breast cancer cells, LINC02086 was consistently found to be up-regulated in breast cancer, and correlated with poorer prognosis. LINC02086 knockdown decreased cell viability, promoted cell apoptosis and suppressed tumor growth. LINC02086 interacted with miR-6757-5p that interacted with EPHA2.LINC02086 expression was negatively correlated with miR-6757-5p expression (r = -0.5698, P < 0.001) but was positively correlated with EPHA2 expression (r = 0.5061, P < 0.001). miR-6757-5p expression was negatively correlated with EPHA2 expression (r = -0.5919, P < 0.001). LINC02086 regulated EPHA2 via miR-6757-5p. miR-6757-5p/EPHA2 axis was a mediator of the effect of LINC02086 on cell viability and apoptosis. CONCLUSION: LINC02086 increases cell viability and decreases apoptotic cells in breast cancer by sponging miR-6757-5p to upregulate EPHA2. This study presents LINC02086/miR-6757-5p/EPHA2 axis as promising therapeutic targets for breast cancer intervention.

Discovery and Characteristics of a Novel Antitumor Cyclopeptide Derived from Shark.

Peptides pose a challenge in drug development due to their short half-lives in vivo. In this study, we conducted in vitro degradation experiments on SAIF, which is a shark-derived peptide that we previously studied. The degradation fragments were sequenced and a truncated peptide sequence was identified. The truncated peptide was then cloned and expressed via the E. coli system with traceless cloning to form a novel cyclic peptide in vitro oxidation condition via the formation of a disulfide bond between the N- and C-termini, which was named ctSAIF. ctSAIF exhibited high anti-HCC activity and enhanced enzymatic stability in vitro, and retained antitumor activity and good biocompatibility in systemic circulation in a HCC xenograft model. Our study discovered and characterized a novel shark-derived cyclic peptide with antitumor activity, laying a foundation for its further development as an antitumor drug candidate. The study also provided a new solution for peptide drug development.

A High Performance Triboelectric Nanogenerator Based on MXene/Graphene Oxide Electrode for Glucose Detection.

A smart sensing platform based on a triboelectric nanogenerator (TENG) possesses various advantages such as self-powering, convenience, real-time and biocompatibility. However, the detection limit of the TENG-based sensor is required to be improved. In this study, a high performance TENG-based glucose sensor was proposed by using the Ti3C2Tx (MXene)/graphene oxide (GO) composite electrode. The MXene and GO nanosheets are popular 2D materials which possessed high conductivity and a rich surface functional group. The MXene/GO thin films were prepared through electrostatic self-assembly technology, which can effectively impede the agglomeration of two nanoflakes. The as-prepared MXene/GO film presented outstanding mechanical property. To figure out the relationship between the nanostructure of MXene/GO film and the TENG, a series of MXene/GO-based TENG with different GO sizes was characterized. As a result, the TENG with 400 nm GO demonstrated the highest output performance. Subsequently, the optimized TENG was used in glucose detection application without the assistance of a glucose enzyme. This simple and flexible TENG shows promising potential in biosensors and non-invasive health monitoring.

A Flexible Triboelectric Nanogenerator Based on Multilayer MXene/Cellulose Nanofibril Composite Film for Patterned Electroluminescence Display.

The flexible self-powered display system integrating a flexible triboelectric nanogenerator (TENG) and flexible alternating current electroluminescence (ACEL) has attracted increasing attention for its promising potential in human-machine interaction applications. In this work, a performance-enhanced MXene/cellulose nanofibril (CNF)/MXene-based TENG (MCM-TENG) is reported for powering a flexible patterned ACEL device in order to realize self-powered display. The MCM multilayer composite film was self-assembled through the layer-by-layer method. The MCM film concurrently acted as a triboelectric layer and electrode layer due to its high conductivity and strength. Moreover, the effect of CNF concentration and number of layers on the output performance of TENG was investigated. It was found that the MCM-TENG realized the optimum output performance. Finally, a flexible self-powered display device was realized by integrating the flexible TENG and ACEL. The MCM-TENG with an output voltage of ≈90 V at a frequency of 2 Hz was found to be efficient enough to power the ACEL device. Therefore, the as-fabricated flexible TENG demonstrates a promising potential in terms of self-powered displays and human-machine interaction.

Alveolar Epithelial Cells Promote IGF-1 Production by Alveolar Macrophages Through TGF-ß to Suppress Endogenous Inflammatory Signals.

To maintain alveolar gas exchange, the alveolar surface has to limit unnecessary inflammatory responses. This involves crosstalk between alveolar epithelial cells (AECs) and alveolar macrophages (AMs) in response to damaging factors. We recently showed that insulin-like growth factor (IGF)-1 regulates the phagocytosis of AECs. AMs secrete IGF-1 into the bronchoalveolar lavage fluid (BALF) in response to inflammatory stimuli. However, whether AECs regulate the production of IGF-1 by AMs in response to inflammatory signals remains unclear, as well as the role of IGF-1 in controlling the alveolar balance in the crosstalk between AMs and AECs under inflammatory conditions. In this study, we demonstrated that IGF-1 was upregulated in BALF and lung tissues of acute lung injury (ALI) mice, and that the increased IGF-1 was mainly derived from AMs. In vitro experiments showed that the production and secretion of IGF-1 by AMs as well as the expression of TGF-ß were increased in LPS-stimulated AEC-conditioned medium (AEC-CM). Pharmacological blocking of TGF-ß in AECs and addition of TGF-ß neutralizing antibody to AEC-CM suggested that this AEC-derived cytokine mediates the increased production and secretion of IGF-1 from AMs. Blocking TGF-ß synthesis or treatment with TGF-ß neutralizing antibody attenuated the increase of IGF-1 in BALF in ALI mice. TGF-ß induced the production of IGF-1 by AMs through the PI3K/Akt signaling pathway. IGF-1 prevented LPS-induced p38 MAPK activation and the expression of the inflammatory factors MCP-1, TNF-α, and IL-1ß in AECs. However, IGF-1 upregulated PPARγ to increase the phagocytosis of apoptotic cells by AECs. Intratracheal instillation of IGF-1 decreased the number of polymorphonuclear neutrophils in BALF of ALI model mice, reduced alveolar congestion and edema, and suppressed inflammatory cell infiltration in lung tissues. These results elucidated a mechanism by which AECs used TGF-ß to regulate IGF-1 production from AMs to attenuate endogenous inflammatory signals during alveolar inflammation.

Clinical characteristics and factors associated with coronary stenosis proximal to a myocardial bridge: a retrospective study.

BACKGROUND: The association of myocardial bridge (MB) with cardiovascular risk and the possible cardiovascular risk factors remain unclear. This study aimed to explore the clinical characteristics and related factors of coronary stenosis proximal to an MB. METHODS: This was a retrospective study of patients with symptoms of coronary atherosclerotic heart disease admitted between 10/2011 and 12/2014 to the Emergency and Cardiology Department of Bayannur Hospital, who underwent selective coronary angiography (SCAG). The patients were assigned to the non-stenosis and stenosis groups according to whether coronary stenosis was proximal to the MB. RESULTS: Among 244 patients with MB and cardiovascular symptoms, 91 (37.3%) had stenosis proximal to the MB. Compared with the non-stenosis group, there were more males (80.2% vs. 55.6%, P < 0.001) and smokers (including those who had quit smoking) (P < 0.001) in the stenosis group. There were no significant differences in blood lipid-related indexes (TG, TC, HDL-C, LDL-C, and VLDL-C) between the two groups. Multivariable analysis suggested that MB location in the middle distal or distal segment of the left anterior descending artery (LAD) increased the odds of coronary stenosis proximal to the MB (OR = 0.439, 95% CI: 1.57-7.532, P = 0.002), which was then considered an independent factor associated with coronary stenosis proximal to the MB. CONCLUSIONS: In patients diagnosed with an MB by SCAG, only MB located in the middle distal or distal segment of the LAD is independently associated with coronary stenosis proximal to the MB.

Crystal structure of the SPRY domain-containing protein 7 reveals unique structural features.

The SPRY/B30.2 domain is one of the most abundant protein domains found in eukaryotes. Vast majority of the SPRY domain-containing proteins are multi-domain proteins. The SPRY domain-containing protein 7 (SPRY7, also named C13orf1, and named chronic lymphocytic leukemia deletion region gene 6 protein, CCLD6, encoded by the spryd7 gene) is the smallest SPRY domain protein in human that does not contain other accessory domains. Here we have determined the crystal structure of human SPRY7 at a resolution of 1.62 Å and found that SPRY7 has some unique structural features that are not present in other previously reported SRPY domain structures. Overall, SPRY7 may represent an evolutionary early version of the SPRY domain, and subsequent loop insertions and expansions, residue substitutions, as well as domain combinations have rendered the SPRY domain versatile binding specificities and broad biological functions. These results serve as a useful basis for a profound characterization of the molecular interactions of SPRY7.

Neutrophil-Lymphocyte Ratio (NLR) for Predicting Clinical Outcomes in Patients with Coronary Artery Disease and Type 2 Diabetes Mellitus: A Propensity Score Matching Analysis.

BACKGROUND: Cardiovascular diseases (CVD) combined with Type 2 diabetes mellitus (T2DM) frequently occurred. In this study, we aimed at exploring the prognostic significance of blood neutrophil-lymphocyte ratio (NLR) in these types of patients. PATIENTS AND METHODS: Between June 30, 2010 and August 30, 2017, 1454 patients with CVD were enrolled in this study. Kaplan and Meier methodology was used for survival analysis. We also used propensity score matching (PSM) to further compare survival in patients with or without T2DM. RESULTS: Among all patients, we applied ROC curve analysis to stratify all patients into two different groups including NLR >2.5 (n=432) and NLR≤ 2.5 (n=1022) groups. After that, we further performed survival analysis between different groups. We found that patients with NLR ≤2.5 had significantly favorable OS compared with the overall survival in patients with NLR >2.5. We further built the PSM using 242 pairs of patients who have CVD and with or without T2DM. After adjusting for competing risk factors, we performed Cox proportional hazards models to identify the independent prognostic factors in multivariable adjustment. We found that NLR ≤2.5 (HR: 2.576, 95% CI: 1.241-4.583, P =0.001) and extent of coronary artery disease (HR: 2.432, 95% CI: 1.189-4.392, P =0.005) remained independent predictors of OS. CONCLUSION: In conclusion, we have established an PSM model and found that a high NLR value was an independent prognostic factor for survival, predicting in patients with both CAD and T2DM. The NLR value would be a valuable biomarker to evaluate the outcomes of patients and give them opportunities for choosing alternative therapies.

[Phagocytosis of alveolar macrophages is suppressed in a mouse model of lipopolysaccharide-induced acute lung injury].

OBJECTIVE: To investigate the changes in phagocytic function of alveolar macrophages (AMs) in mice with lipopolysaccharide (LPS)-induced acute lung injury (ALI) and explore the possible mechanism. METHODS: Kunming mice were randomly divided into normal control group and ALI (induced by LPS instillation in the airway) model group. AMs were obtained from bronchoalveolar lavage fluid in both groups, and phagocytosis of the AMs was observed using flow cytometry and fluorescence microscopy. Western blotting and ELISA were used to detect the expression and secretion of IL-33 in the lung tissue of the mice. We also detected the secretion of IL-33 by an alveolar epithelial cell line MLE-12 in response to stimulation with different concentrations of LPS. The AMs from the normal control mice were treated with different concentrations of LPS and IL-33, and the changes in the phagocytic activity of the cells were observed. RESULTS: Compared with those in normal control group, the percentage of AMs phagocytosing fluorescent microspheres was significantly decreased, and the expression of IL-33 in lung tissue and IL-33 level in the bronchoalveolar lavage fluid were significantly increased in ALI mice (P < 0.01). LPS (100-1000 ng/mL) obviously promoted the secretion of IL-33 in cultured MLE-12 cells (P < 0.01). Both LPS (10-500 ng/mL) and IL-33 (100 ng/mL) significantly inhibited the phagocytic activity of the AMs from normal control mice (P < 0.01). CONCLUSIONS: The phagocytic activity of AMs is weakened in ALI mice possibly due to direct LPS stimulation and the inhibitory effect of the alarmin IL-33 produced by LPS-stimulated alveolar epithelial cells.

SCF promotes the production of IL-13 via the MEK-ERK-CREB signaling pathway in mast cells.

Mast cells serve a key role in the occurrence and development of allergy. As an important growth factor of mast cells, stem cell factor (SCF) has an effect on the apoptosis, chemotaxis, adhesion, degranulation and other biological characteristics of mast cells. However, there are few studies regarding the effect of SCF signal on the production of cytokines from mast cells, particularly Th2 type cytokines. In the present study, the expression and secretion of IL-13 in P815 cells stimulated by SCF were detected by fluorescence quantitative PCR and ELISA, and western blotting and EMSA were used to detect ERK phosphorylation and activation of CREB in stimulated P815 cells. The results demonstrated that the production of IL-13 was significantly increased in P815 cells stimulated by SCF (1-100 ng/ml; P<0.01). There was an obvious phosphorylation of ERK and CREB activation in P815 cells stimulated by SCF (50 ng/ml). Compared with the SCF single stimulation group, the production of IL-13 was significantly reduced in P815 cells stimulated with U0126 (ERK-MEK/pathway inhibitor) or H-89 (CREB inhibitor) combined with SCF stimulation group (P<0.01). However, JSI-124 (JAK/STAT3 pathway inhibitor), Wortmannin (PI3K/Akt pathway inhibitor) and PDTC (NF-κB inhibitor) had no effect on the role of SCF promoting the P815 cells producing IL-13. Therefore, SCF signaling promotes mast cell P815 to produce IL-13, and this effect is associated with the MEK-ERK-CREB signaling pathway.

Insulin­like growth factor 1 inhibits phagocytosis of alveolar epithelial cells in asthmatic mice.

The phagocytosis of apoptotic cells by alveolar epithelial cells helps to eliminate airway inflammation. Insulin­like growth factor 1 (IGF­1) regulates cell metabolism and proliferation, and promotes cell survival, while it may also promote the proliferation and differentiation of alveolar epithelial cells during the repair of lung injury. The present study investigated the effect of IGF­1 on the phagocytic activity of alveolar epithelial cells, a nonprofessional phagocyte. IGF­1 was elevated in lung tissue and bronchoalveolar lavage fluid obtained from mice with ovalbumin­induced asthma. IGF­1 was reduced by 50% in the lung tissue and by nearly 100% in the bronchoalveolar lavage fluid in asthmatic mice established by depletion of alveolar macrophages using 2­chloroadenosine. In addition, interleukin­33 induced IGF­1 production in primary alveolar macrophages. It was also observed that IGF­1 inhibited the phagocytosis of fluorescent microspheres and apoptotic cells by MLE­12 alveolar epithelial cells. Antibody blocking of IGF­1 enhanced the phagocytosis of fluorescent microspheres and apoptotic cells, and significantly reduced inflammatory cell infiltration in airway and perivascular tissues. The elevated IGF­1 level in the lungs of asthma model mice was mainly produced in alveolar macrophages. Taken together, the current study demonstrated that IGF­1 inhibited phagocytosis by alveolar epithelial cells, and that IGF­1 blockade enhanced the phagocytic activity and alleviated airway inflammation. These results support the potential use of IGF­1 as a target in the treatment of asthma.

Backbone 1H, 13C, and 15N resonance assignments of the PRY-SPRY domain of RNF135.

RING finger protein 135 (RNF135, also named Riplet or REUL) exerts multiple biological functions and its C-terminal PRY-SPRY/B30.2 domain is indispensable for most of these functions. RNF135 interacts with RIG-I (retinoic acid-inducible gene-I) via the PRY-SPRY domain and ubiquitinates RIG-I to promote innate anti-viral signaling, while mutations in the RNF135 gene can cause the Macrocephaly, macrosomia, facial dysmorphism (MMFD) syndrome, and RNF135 reportedly regulates the proliferation of glioblastoma cells as well as tongue cancer cells. Nevertheless, structure of full-length RNF135 or its PRY-SPRY domain has not been determined, and structural basis for molecular interactions involving RNF135 is largely unknown. Here we report the backbone 1H, 13C, and 15N chemical shift assignments of the PRY-SPRY domain of RNF135 and the secondary structure elements predicted based on chemical shifts, as well as the perturbations caused by the R286H mutation that is associated with MMFD syndrome. We found that the mutation did not alter the gross structure of the PRY-SPRY domain, so it may have impaired RNF135 function by affecting protein-protein interactions mediated by the domain.

MicroRNA-20b promotes proliferation of H22 hepatocellular carcinoma cells by targeting PTEN.

MicroRNAs (miRNAs/miRs) are small, noncoding RNA molecules that are closely associated with the occurrence and development of tumors. miR-20b is overexpressed in hepatocellular carcinoma cell lines and tissues. However, it is not clear whether miR-20b can promote the proliferation of hepatocellular carcinoma cells. In the present study, the proliferation of H22 mouse hepatocellular carcinoma cells was detected using the Cell Counting Kit-8 assay. MiRanda software was used to predict the binding sites of miR-20b to the 3'-untranslated region (3'-UTR) of phosphatase and tensin homolog (PTEN). The 3'-UTR sequence of the PTEN gene was amplified using the polymerase chain reaction in H22 cells. The recombinant plasmid or empty plasmid was co-transfected with miR-20b mimics or miR-20b scramble into HeLa cells, and luciferase activity was assessed by Dual-Luciferase® Reporter Assay System 24 h post-transfection. In the present study, miR-20b knockdown significantly inhibited the proliferation of H22 mouse hepatocellular carcinoma cells. In addition, miR-20b inhibition upregulated the expression of PTEN, and it was revealed that miR-20b may directly target the 3'-untranslated region of the PTEN gene. Downregulation of PTEN partially reversed the anti-proliferative effect of miR-20b on H22 cells. In conclusion, miR-20b may promote H22 cell proliferation by targeting PTEN, providing a rationale for further study investigating novel therapeutic strategies for liver cancer.

IGF-1 Promotes Endocytosis of Alveolar Epithelial Cells through PI3K Signaling.

Airway inflammation can be mitigated when apoptotic cells are engulfed by pulmonary epithelial cells. Insulin-like growth factor 1 (IGF-1), a single chain polypeptide growth factor, is the main mediator of growth hormone activity in vivo. IGF-1 has many biological activities, such as the regulation of cell survival, proliferation, differentiation and metabolism. However, its effect on the engulfment of cells, especially by non-professional phagocytes such as alveolar epithelial cells (AECs), has not been fully elucidated. We report that IGF-1 increases endocytosis in a mouse alveolar epithelial cell line, MLE-12. The PI3K-Akt pathway is involved in this effect of IGF-1. Furthermore, IGF-1 can inhibit the production of interleukin-6 in lipopolysaccharide-stimulated AECs. We have found that IGF-1 can enhance endocytosis of AECs through the PI3K pathway and exhibit anti-inflammatory properties. These two observations suggest that IGF-1 is a potential mediator in the regulation of airway inflammation.

Magnetic liposomes for light-sensitive drug delivery and combined photothermal-chemotherapy of tumors.

The targeted delivery of anticancer drugs for improving the therapeutic efficacy and reducing the side effects has attracted great attention in cancer therapy. In this study, multifunctional magnetic nanoparticles-loaded thermosensitive liposomes (Fe3O4-TSL) were developed for the near-infrared (NIR) laser-triggered release and combined photothermal-chemotherapy of tumors. Doxorubicin (DOX) was encapsulated into the Fe3O4-TSL (DOX-Fe3O4-TSL) via an ammonium sulfate gradient, with an encapsulation efficiency of up to 90.9%. Once treated with NIR laser irradiation, significantly improved drug release was observed in the DOX-Fe3O4-TSL compared to that of DOX-TSL. After an intravenous injection, Fe3O4-TSL tended to enrich in the tumor over time and showed remarkable magnetic resonance imaging (MRI) and photothermal effects. The combined chemo-photothermal therapy study demonstrated that DOX-Fe3O4-TSL could significantly inhibit the tumor growth without causing any significant damage to normal tissues under NIR laser irradiation. These results revealed a great potential for DOX-Fe3O4-TSL for the diagnosis and synergistic therapy of tumors.

Upregulated IGF­1 in the lungs of asthmatic mice originates from alveolar macrophages.

Asthma is characterized by inflammation and remodeling of the airways. Insulin­like growth factor-1 (IGF­1) serves an important role in the repair of lung tissue injury and airway remodeling by elevating collagen and elastin content, increasing the thickness of smooth muscle and promoting the proliferation of lung epithelial and interstitial cells, as well as fibroblasts; however, the content of IGF­1 and its cellular origin in the lungs of patients with asthma remain unknown. In the present study, a mouse model of asthma was constructed. Following isolation of alveolar macrophages (AMs), the content of IGF­1 in lung tissue and bronchoalveolar lavage fluid (BALF) was detected by ELISA. The proliferation and phagocytosis of alveolar epithelial cells (AECs) stimulated by IGF­1 were detected by Cell Counting Kit­8 method and flow cytometry, respectively. In the present study, IGF­1 was upregulated in the lung tissues of asthmatic mice, and the content of IGF­1 in BALF was also elevated. Depletion of AMs by treating mice with 2­chloroadenosine via nose dripping reversed the increase of IGF­1 by 80% in lung tissues and by ~100% in BALF of asthmatic mice, suggesting that elevated IGF­1 in asthmatic mice predominantly originated from AMs. As IGF­1 promotes the proliferation and phagocytosis of AECs, AM­derived IGF­1 may serve an important role in the regulation of airway inflammation and remodeling in asthmatic mice.

miR­20b negatively regulates VEGF expression by targeting STAT3 in H22 hepatocellular carcinoma cells.

Vascular endothelial growth factor (VEGF) promotes angiogenesis during tumor growth, and its expression involves multiple signaling pathways and transcription factors. In the present study, transforming growth factor (TGF)­ß1 promoted upregulation of VEGF and downregulation of microRNA (miR)­20b expression in mouse H22 hepatocellular carcinoma cells. miR­20b negatively regulated both constitutive VEGF expression and TGF­ß1­induced VEGF expression. The miRanda algorithm predicted that a binding site of the miR­20b GCAAUCUGGGCACUUU sequence was present in the signal transducer and activator of transcription (STAT)3 3'­untranslated region. Following transfection of miR­20b mimics into H22 cells, expression of STAT3 protein was downregulated. A dual­luciferase activity assay revealed that miR­20b directly targeted STAT3 to regulate its expression, and that interference with STAT3 expression significantly downregulated VEGF mRNA and protein expression. Interference with STAT3 expression resulted in increased VEGF expression in H22 cells overexpressing miR­20b, but expression was lower than that in quiescent H22 cells. This indicated that STAT3 was involved in the negative regulation of VEGF expression in H22 cells by miR­20b. The data demonstrated that miR­20b negatively regulated VEGF expression by directly targeting STAT3 in H22 cells.

Reduced peripheral blood regulatory B cell levels are not associated with the Expanded Disability Status Scale score in multiple sclerosis.

Objective To investigate levels of regulatory B (Breg) cells, plasma cells, and memory B cells in the peripheral blood, and interleukin (IL)-10 in the serum of multiple sclerosis (MS) patients, and to determine the correlation between Breg cell levels and the Expanded Disability Status Scale (EDSS) score. Methods Levels of Breg cells, plasma cells, and memory B cells in the peripheral blood of 12 MS patients were measured using flow cytometry. IL-10 serum levels were measured by enzyme-linked immunosorbent assay. The correlation between Breg cell levels and MS EDSS score was measured using Pearson's correlation coefficient. Results Compared with healthy controls, MS patients had decreased levels of CD19+CD24hiCD38hi Breg cells in their peripheral blood and reduced serum levels of IL-10; however, the ratios of CD19+CD27hiCD38hi plasma cells and CD19+CD27+CD24hi memory B cells to total B cells did not differ significantly between healthy controls and MS patients. CD19+CD24hiCD38hi Breg cell levels in the peripheral blood of MS patients were not significantly correlated with MS EDSS score. Conclusion Peripheral blood CD19+CD24hiCD38hi Breg cell levels and serum IL-10 levels were reduced in MS patients compared with controls, but Breg cell levels were not correlated with MS EDSS score.